New CRISPR System For Gene Silencing Avoids DNA Cuts

Scientists at Vilnius University's Life Sciences Center (VU-LSC) have discovered a novel method for cells to silence specific genes without cutting DNA.

The research team uncovered how cells can use a specialized system to locate and silence unwanted DNA. I Photo: Vilnius University

Led by Prof. Patrick Pausch and published in Nature Communications, this research introduces a way to silence genes by effectively "pausing" certain genetic instructions within cells.

The research team, including doctoral student Rimvydė Čepaitė, Dr. Aistė Skorupskaitė, undergraduate Gintarė Žvejyte, and Prof. Pausch, along with an international team, uncovered how cells can use a specialized system to locate and silence unwanted DNA.

This approach could lead to safer gene modifications, potentially offering new treatments for diseases caused by faulty genes.

"Unlike the well-known CRISPR gene-editing system, often described as molecular 'scissors,' this newly studied type IV-A CRISPR system doesn’t cut genes. Instead, it uses an RNA-guided 'effector' complex to recruit an enzyme called DinG, which travels along DNA to silence targeted genes more subtly," explains Prof. Pausch.

The system’s ability to accurately identify the specific DNA location where it will function is particularly remarkable.

The system uses two proteins, Cas8 and Cas5, to find a short sequence motif adjacent to the RNA guide’s complementary target DNA. Once both proteins recognize this sequence, they open the double-stranded DNA for further targeting.

A key step is the formation of R-loops—open DNA structures where RNA binds, signaling the system to begin silencing the gene.

"The 'R' in R-loop stands for RNA. All DNA-binding CRISPR-Cas systems rely on this structure to verify DNA sequences and locate the correct target site. Stable R-loops only form in the presence of a DNA sequence that matches the guide RNA, essentially giving the system a 'green light' to silence the gene," explains Prof. Pausch.

The DinG enzyme enhances gene suppression by unwinding DNA strands, allowing the system to extend its effect along a longer DNA sequence.